Role of Fibulin-1 in APP Processing

描述（由申请人提供）：目前许多对抗阿尔茨海默病（AD）的药理学方法试图通过抑制淀粉样蛋白生成酶（称为β-和γ-分泌酶）来阻断Abeta的产生。另一种方法是激活淀粉样前体蛋白（APP）的α-分泌酶加工，这是由金属蛋白酶的去整合素家族的几个成员，ADAM 9，ADAM 10和ADAM 17介导的。通过这些α-分泌酶加工APP被认为对于AD是有益的，因为它限制了Abeta的产生并产生神经保护性可溶性APP α（sAPP α）产物。Fibulin-1（Fbln 1）是一种细胞外基质蛋白，由神经元在脑中表达，其结合APP和sAPP α的氨基末端。这种相互作用的意义尚未确定，然而，我们发现Fbln 1也结合其他膜锚定α-分泌酶底物，肝素结合表皮生长因子（HB-EGF）和神经调节蛋白-1（NRG 1）。我们还表明，Fbln 1的行为，以抑制HB-EGF和NRG 1的可溶性形式的蛋白水解释放。此外，我们已经发现与野生型MEF相比，在Fbln 1缺失小鼠胚胎成纤维细胞（MEF）的条件培养基中sAPP α的水平增加。基于这些发现，假设Fbln 1作为APP的α-分泌酶加工的抑制剂，因此可能代表治疗靶点，如果被抑制，可能导致APP的α-分泌酶加工增强和病理性APP裂解减少。为了解决这一假设，有三个具体的目标：1）确定脑APP蛋白水解裂解是否改变Fbln 1缺陷小鼠，2）确定是否转基因Fbln 1过表达加速A β生产和加剧AD发病机制，和3）确定是否Fbln 1抑制α分泌酶加工的APP在培养的神经元细胞。

DESCRIPTION (provided by applicant): Many current pharmacological approaches to combating Alzheimer's disease (AD) seek to block Abeta production through inhibition of the amyloidogenic enzymes known as beta- and gamma-secretases. An alternative approach is to activate the alpha-secretase processing of amyloid precursor protein (APP), which is mediated by several members of the disintegrin family of metalloproteases, ADAM9, ADAM10 and ADAM17. Processing of APP by these alpha-secretases is thought to be beneficial with respect to AD since it limits production of Abeta and generates the neuroprotective soluble APPalpha (sAPPalpha) product. Fibulin-1 (Fbln1) is an extracellular matrix protein, expressed in the brain by neurons, that binds the amino terminus of APP and sAPPalpha. The significance of this interaction is not yet established however, we have found that Fbln1 also binds to other membrane anchored alpha-secretase substrates, heparin binding-epidermal growth factor (HB-EGF) and neuregulin-1 (NRG1). We also show that Fbln1 acts to inhibit the proteolytic release of soluble forms of HB-EGF and NRG1. Furthermore, we have found increased levels of sAPPalpha in the conditioned culture medium of Fbln1 null mouse embryo fibroblasts (MEFs) as compared to wildtype MEFs. Based on these findings it is hypothesized that Fbln1 serves as an inhibitor of alpha-secretase processing of APP and therefore may represent a therapeutic target that if inhibited might lead to augmented alpha-secretase processing of APP and reduced pathological APP cleavage. To address this hypothesis there are three specific aims: 1) Determine whether brain APP proteolytic cleavage is altered in Fbln1-deficient mice, 2) determine whether transgenic overexpression of Fbln1 accelerates Abeta production and exacerbates AD pathogenesis, and 3) determine whether Fbln1 inhibits alpha-secretase processing of APP in cultured neuronal cells.